Prolyl isomerase Pin1 regulates cadmium-induced autophagy via ubiquitin-mediated post-translational stabilization of phospho-Ser GSK3αβ in human hepatocellular carcinoma cells

Low Cd (≤IC50) exposure transiently increased p-Ser GSK3αβ and polyubiquitinated protein levels, but only high Cd concentrations decreased Pin1 levels, and constitutively accumulated p-Ser GSK3αβ and poly-ubiquitinated protein, and increased autophagy and apoptosis. [Display omitted] Accumulating evidence suggests that prolyl-isomerase (Pin1) is an important regulator of apoptosis. In the present study, Pin1 was shown to negatively regulate the stabilization of glycogen synthase kinase (GSK) 3αβ serine phosphorylation by inhibiting ubiquitin (Ub)-proteasome degradation, and to induce autophagy following cadmium (Cd) exposure. Cd-induced autophagy in human hepatoma (HepG2) cells has been demonstrated by green fluorescent protein (GFP)-LC3B plasmid DNA transfection, and LC3-II conversion by autophagy inhibitors. Atg5 silencing inhibited Cd-induced apoptosis, indicating that autophagy is involved in cell death. Exposing HepG2 cells to Cd (≤6μM) initially increased p-Ser GSK3αβ, but then resulted in a gradual decrease, which correlated with polyubiquitinated protein levels, which is indicative of protein degradation. However, high Cd concentrations lead to p-Ser GSK3αβ and polyubiquitinated protein accumulation, indicating proteasome impairment. Cd-induced p-Ser GSK3αβ was enhanced by proteasome inhibition (MG132) and ubiquitin deficiency (cyclohexamide), but no ubiquitination of p-Ser GSK3αβ or GSK3αβ was detected. Cd exposure resulted in p-Ser GSK3αβ redistribution from the nucleus into the cytosol, and this along with autophagy, was inhibited by leptomycin B. Cd concentrations